Normal cardiovascular adaptations in pregnancy include generalized vasodilation and reduced uterine and systemic vasoconstriction in response to infused angiotensin II (ANG II) and alpha-agonists. During human and ovine pregnancy the uterine vascular bed is even less reactive to vasoconstriction by ANG II compared to systemic vasculature. The long-term goals of this research project are to determine if the vasodilatory endothelial factors prostacyclin (PGI2) and endothelium- derived relaxing factor/nitric oxide (EDRF/NO) are responsible for modulating maternal vascular smooth muscle (VSM) function in normal pregnancy and if estrogen and/or progesterone, steroid hormones elevated in pregnancy, will modulate their productions. We know from in vitro models which evaluate endothelial/VSM interactions that endothelium- derived PGI2 and EDRF/NO are released in a paracrine fashion to modulate production of the second messenger cyclic nucleotides cAMP and cGMP, respectively, which then decrease VSM constriction and tone. The overall hypothesis of this project is that uterine and systemic endothelium- derived PGI2 and EDRF/NO productions are increased in normal pregnancy and that they modulate VSM cAMP and cGMP to control dilation and arterial responsiveness to vasoconstrictors in particular ANG II. The specific aims of this project are to define the molecular and enzymatic mechanisms controlling PGI2 and EDRF/NO production in normal pregnancy in order to further understand uterine and systemic artery endothelial modulation of VSM cAMP and cGMP production. We will evaluate adenylate and guanylate cyclase enzyme sensitivity in VSM from pregnant and nonpregnant ewes. Also planned is the delineation of which enzymes are activated in association with increased production of endothelium-derived PGI2 and EDRF/NO during pregnancy; cyclooxygenase (PGHS-1 & PGHS-2), PGI2 synthase and EDRF/NO synthase will be studied using Western immunoblots, immunocytochemistry and productions of PGI2, cAMP, and cGMP; specific activity studies also are planned. Experiments are proposed to study the gene regulation of cyclooxygenase, PGI2 synthase EDRF/NO synthase in pregnancy using Northern blots and in situ hybridization. The direct effects of estrogen and/or progesterone to regulate PGI2 and EDRF/NO production and gene expression will provide mechanisms by which placental/ovarian steroid production modifies VSM function in pregnancy. Addressing these aims will increase our knowledge concerning the enzymatic and molecular control endothelial/VSM interactions associated with normal and potentially pathologic cardiovascular adaptations in pregnancy.